Chapelet Guillaume, Béguin Nora, Castellano Blandine, Grit Isabelle, de Coppet Pierre, Oullier Thibauld, Neunlist Michel, Blottière Hervé, Rolli-Derkinderen Malvyne, Le Dréan Gwenola, Derkinderen Pascal
Nantes Université, INSERM, CHU Nantes, The Enteric Nervous System in Gut and Brain Disorders, Nantes, France.
Nantes Université, INRAE, IMAD, CRNH-O, UMR 1280, PhAN, Nantes, France.
Front Neurosci. 2023 Jun 2;17:1166848. doi: 10.3389/fnins.2023.1166848. eCollection 2023.
There is mounting evidence to suggest that the gut-brain axis is involved in the development of Parkinson's disease (PD). In this regard, the enteroendocrine cells (EEC), which faces the gut lumen and are connected with both enteric neurons and glial cells have received growing attention. The recent observation showing that these cells express alpha-synuclein, a presynaptic neuronal protein genetically and neuropathologically linked to PD came to reinforce the assumption that EEC might be a key component of the neural circuit between the gut lumen and the brain for the bottom-up propagation of PD pathology. Besides alpha-synuclein, tau is another key protein involved in neurodegeneration and converging evidences indicate that there is an interplay between these two proteins at both molecular and pathological levels. There are no existing studies on tau in EEC and therefore we set out to examine the isoform profile and phosphorylation state of tau in these cells.
Surgical specimens of human colon from control subjects were analyzed by immunohistochemistry using a panel of anti-tau antibodies together with chromogranin A and Glucagon-like peptide-1 (two EEC markers) antibodies. To investigate tau expression further, two EEC lines, namely GLUTag and NCI-H716 were analyzed by Western blot with pan-tau and tau isoform specific antibodies and by RT-PCR. Lambda phosphatase treatment was used to study tau phosphorylation in both cell lines. Eventually, GLUTag were treated with propionate and butyrate, two short chain fatty acids known to sense EEC, and analyzed at different time points by Western blot with an antibody specific for tau phosphorylated at Thr205.
We found that tau is expressed and phosphorylated in EEC in adult human colon and that both EEC lines mainly express two tau isoforms that are phosphorylated under basal condition. Both propionate and butyrate regulated tau phosphorylation state by decreasing its phosphorylation at Thr205.
Our study is the first to characterize tau in human EEC and in EEC lines. As a whole, our findings provide a basis to unravel the functions of tau in EEC and to further investigate the possibility of pathological changes in tauopathies and synucleinopathies.
越来越多的证据表明，肠-脑轴参与帕金森病（PD）的发病过程。在这方面，面向肠腔且与肠神经元和神经胶质细胞均相连的肠内分泌细胞（EEC）受到了越来越多的关注。最近的观察表明，这些细胞表达α-突触核蛋白，一种在基因和神经病理学上与PD相关的突触前神经元蛋白，这进一步强化了一种假设，即EEC可能是肠腔与大脑之间神经回路的关键组成部分，参与PD病理的自下而上传播。除了α-突触核蛋白，tau是另一种参与神经退行性变的关键蛋白，越来越多的证据表明这两种蛋白在分子和病理水平上存在相互作用。目前尚无关于EEC中tau的研究，因此我们着手研究这些细胞中tau的异构体谱和磷酸化状态。
使用一组抗tau抗体以及嗜铬粒蛋白A和胰高血糖素样肽-1（两种EEC标志物）抗体，通过免疫组织化学分析来自对照受试者的人类结肠手术标本。为了进一步研究tau的表达，使用泛tau和tau异构体特异性抗体通过蛋白质印迹法以及逆转录聚合酶链反应（RT-PCR）分析两种EEC细胞系，即GLUTag和NCI-H716。使用λ磷酸酶处理来研究两种细胞系中的tau磷酸化。最后，用丙酸盐和丁酸盐处理GLUTag，这两种短链脂肪酸已知可作用于EEC，并在不同时间点用针对苏氨酸205磷酸化的tau的特异性抗体通过蛋白质印迹法进行分析。
我们发现tau在成人人类结肠的EEC中表达并磷酸化，并且两种EEC细胞系主要表达两种在基础条件下磷酸化的tau异构体。丙酸盐和丁酸盐均通过降低其在苏氨酸205处的磷酸化来调节tau的磷酸化状态。
我们的研究首次对人类EEC和EEC细胞系中的tau进行了表征。总体而言，我们的研究结果为揭示tau在EEC中的功能以及进一步研究tau蛋白病和突触核蛋白病中病理变化的可能性提供了依据。
